Definition of in vitro model systems in which non MHC-restricted cytotoxic cells are generated under conditions of lymphocytes' stimulation constitutes an essential tool for the understanding of the lineage and biology of the cytotoxic cells, of the factors regulating their proliferation and activation, and of the interplay occurring in vivo in pathological situations among different cell types effector of the immune and non immune system. We developed an in vitro system in which high numbers of cytotoxic cells are generated, depending on endogenous IL-2 production, from peripheral blood mononuclear cells cocultured with B lymphoblastoid lines, even if these lack class I HLA antigens. Most, but not all, of these expanded cytotoxic lymphocytes have characteristics of NK cells. Moreover, we observed that B lymphoblastoid lines constitutively produce a factor that potentiates several functional properties of NK cells, among which are cytotoxic activity, production of lymphokines and proliferation. This factor is clearly distinct from other known lymphokines such as IL-2, interferon (IFN) or lymphotoxin (LT). It is our working hypothesis that, in vivo, proliferation, differentiation and functions of NK cells and of other non well defined cytotoxic cell types (among which possibly LAK cells) are regulated not only by IL-2, but also by other cytokines, possibly produced by accessory cells or by the NK cells themselves, upon cellular interaction or IL-2 stimulation. The system we developed and our observation that NKSF, a factor distinct from IL-2, activates NK cells offer the opportunity to study the regulation of NK cell activation and proliferation under conditions similar to those possibly present in pathological in vivo situations in which minimal antigenic differences may induce lymphocyte activation. The studies proposed to characterize our in vitro model and NKSF should provide information on NK cells' biology and on the practical possibility of enhancing efficiently their number and functions.